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The All-in-One digital microfluidic pipeline allows for end-to-end automation of processing and analysis of mass-limited proteomic samples.

Microfluidic platforms are an attractive option for incorporating complex fluid handling into low-cost and rapid diagnostic tests. A persistent challenge for microfluidics, however, is the mismatch in the "world-tochip" interface - it is challenging to detect analytes present at low concentrations in systems that can only handle small volumes of sample. Here we describe a new technique termed pre-concentration by liquid intake by paper (P-CLIP) that addresses this mismatch, allowing digital microfluidics to interface with volumes on the order of hundreds of microliters. In P-CLIP, a virtual microchannel is generated to pass a large volume through the device; analytes captured on magnetic particles can be isolated and then resuspended into smaller volumes for further processing and analysis. We characterize this method and demonstrate its utility with an immunoassay for Plasmodium falciparum lactate dehydrogenase, a malaria biomarker, and propose that the P-CLIP strategy may be useful for a wide range of applications that are currently limited by low-abundance analytes.

We studied the effect of AHR expression on metastasis using cell invasion in digital microfluidic microgel systems (CIMMS), which provided a unique combination of functional discrimination with transcriptome profiling of sub-populations of cells.

The rapid development of micromanipulation technologies has opened exciting new opportunities for the actuation, selection and assembly of a variety of non-biological and biological nano/micro-objects for applications ranging from microfabrication, cell analysis, tissue engineering, biochemical sensing, to nano/micro-machines. To date, a variety of precise, flexible and high-throughput manipulation techniques have been developed based on different physical fields. Among them, optoelectronic tweezers (OET) is a state-of-art technique that combines light stimuli with electric field together by leveraging the photoconductive effect of semiconductor materials. Herein, the behavior of micro-objects can be directly controlled by inducing the change of electric fields on demand in an optical manner. Relying on this light-induced electrokinetic effect, OET offers tremendous advantages in micromanipulation such as programmability, flexibility, versatility, high-throughput and ease of integration with other characterization systems, thus showing impressive performance compared to those of many other manipulation techniques. A lot of research on OET have been reported in recent years and the technology has developed rapidly in various fields of science and engineering. This work provides a comprehensive review of the OET technology, including its working mechanisms, experimental setups, applications in non-biological and biological scenarios, technology commercialization and future perspectives.

Supercritical angle fluorescence (SAF) microscopy is a novel imaging tool based on the use of distance-dependent fluorophore emission patterns to provide accurate locations of fluorophores relative to a surface. This technique has been extensively used to construct accurate cellular images and to detect surface phenomena in a static environment. However, the capability of SAF microscopy in monitoring dynamic surface phenomena and changes in millisecond intervals is underexplored in the literature. Here, we report on a hardware add-on for a conventional inverted microscope coupled with a post-processing Python module that extends the capability of SAF microscopy to monitor dynamic surface adsorption in sub-second intervals, thereby greatly expanding the potential of this tool to study surface interactions, such as surface fouling and competitive surface adhesion. The Python module enables researchers to automatically extract SAF profiles from each image. We first assessed the performance of the system by probing the specific binding of biotin-fluorescein conjugates to a neutravidin-coated cover glass in the presence of non-binding fluorescein. The SAF emission was observed to increase with the quantity of bound fluorophore on the cover glass. However, a high concentration of unbound fluorophore also contributed to overall SAF emission, leading to over-estimation in surface-bound fluorescence. To expand the applications of SAF in monitoring surface phenomena, we monitored the non-specific surface adsorption of BSA and non-ionic surfactants on a Teflon-AF surface. Solution mixtures of bovine serum albumin (BSA) and nine Pluronic/Tetronic surfactants were exposed to a Teflon-AF surface. No significant BSA adsorption was observed in all BSA-surfactant solution mixtures with negligible SAF intensity. Finally, we monitored the adsorption dynamics of BSA onto the Teflon-AF surface and observed rapid BSA adsorption on Teflon-AF surface within 10 s of addition. The adsorption rate constant (ka) and half-life of BSA adsorption on Teflon-AF were determined to be 0.419 ± 0.004 s−1 and 1.65 ± 0.016 s, respectively, using a pseudo-first-order adsorption equation.

Aaron R. Wheeler and Philippa Ross introduce the first Lab on a Chip Reviews issue.

Bacteria are widely studied in various research areas, including synthetic biology, sequencing and diagnostic testing. Protocols involving bacteria are often multistep, cumbersome and require access to a long list of instruments to perform experiments. In order to streamline these processes, the fluid handling technique digital microfluidics (DMF) has provided a miniaturized platform to perform various steps of bacterial protocols from sample preparation to analysis. DMF devices can be paired/interfaced with instrumentation such as microscopes, plate readers, and incubators, demonstrating their versatility with existing research tools. Alternatively, DMF instruments can be integrated into all-in-one packages with on-chip magnetic separation for sample preparation, heating/cooling modules to perform assay steps and cameras for absorbance and/or fluorescence measurements. This perspective outlines the beneficial features DMF offers to bacterial protocols, highlights limitations of current work and proposes future directions for this tool's expansion in the field.

Optoelectronic tweezers (OET) is a noncontact micromanipulation technology for controlling microparticles and cells. In the OET, it is necessary to configure a medium with different electrical properties to manipulate different particles and to avoid the interaction between two particles. Here, a new method exploiting the interaction between different dielectric properties of micro-objects to achieve the trapping, transport, and release of particles in the OET system was proposed. Besides, the effect of interaction between the micro-objects with positive and negative dielectric properties was simulated by the arbitrary Lagrangian-Eulerian (ALE) method. In addition, compared with conventional OET systems relying on fabrication processes involving the assembly of photoelectric materials, a contactless OET platform with an iPad-based wireless-control interface was established to achieve convenient control. Finally, this platform was used in the interaction of swimming microorganisms (positive-dielectric properties) with microparticles (negative-dielectric properties) at different scales. It showed that one particle could interact with 5 particles simultaneously, indicating that the interaction can be applied to enhance the high-throughput transportation capacities of the OET system and assemble some special microstructures. Owing to the low power, microorganisms were free from adverse influence during the experiment. In the future, the interaction of particles in a simple OET platform is a promising alternative in micro-nano manipulation for controlling drug release from uncontaminated cells in targeted therapy research.

We introduce Digital microfluidic Isolation of Single Cells for -Omics (DISCO), a platform that allows users to select particular cells of interest from a limited initial sample size and connects single-cell sequencing data to their immunofluorescence-based phenotypes. Specifically, DISCO combines digital microfluidics, laser cell lysis, and artificial intelligence-driven image processing to collect the contents of single cells from heterogeneous populations, followed by analysis of single-cell genomes and transcriptomes by next-generation sequencing, and proteomes by nanoflow liquid chromatography and tandem mass spectrometry. The results described herein confirm the utility of DISCO for sequencing at levels that are equivalent to or enhanced relative to the state of the art, capable of identifying features at the level of single nucleotide variations. The unique levels of selectivity, context, and accountability of DISCO suggest potential utility for deep analysis of any rare cell population with contextual dependencies., Multi-Omic approaches are a powerful way for obtaining in-depth understanding of a cell’s state. Here the authors present DISCO, combining digital microfluidics, laser cell lysis, and artificial intelligence-driven image processing to analyze single-cell genomes, transcriptomes and proteomes in a mixed population.

Finger-stick blood sampling is convenient for point of care diagnostics, but whole blood samples are problematic for many assays because of severe matrix effects associated with blood cells and cell debris. We introduce a new digital microfluidic (DMF) diagnostic platform with integrated porous membranes for blood-plasma separation from finger-stick blood volumes, capable of performing complex, multi-step, diagnostic assays. Importantly, the samples can be directly loaded onto the device by a finger "dab" for user-friendly operation. We characterize the platform by comparison to plasma generated via the "gold standard" centrifugation technique, and demonstrate a 21-step rubella virus (RV) IgG immunoassay yielding a detection limit of 1.9 IU mL-1, below the diagnostic cut-off. We propose that this work represents a critical next step in DMF based portable diagnostic assays-allowing the analysis of whole blood samples without pre-processing.

High-throughput fluidic technologies have increased the speed and accuracy of fluid processing to the extent that unlocking further gains will require replacing the human operator with a robotic counterpart. Recent advances in chemistry and biology, such as gene editing, have further exacerbated the need for smart, high-throughput experimentation. A growing number of innovations at the intersection of robotics and fluidics illustrate the tremendous opportunity in achieving fully self-driving fluid systems. We envision that the fields of synthetic chemistry and synthetic biology will be the first beneficiaries of AI-directed robotic and fluidic systems, and largely fall within two modalities: complex integrated centralized facilities that produce data, and distributed systems that synthesize products and conduct disease surveillance.

Correction for ‘Virtual microwells for digital microfluidic reagent dispensing and cell culture’ by Irwin A. Eydelnant et al., Lab Chip, 2012, 12, 750–757, https://doi.org/10.1039/C2LC21004E.

With the prevalence of bacterial infections and increasing levels of antibiotic resistance comes the need for rapid and accurate methods for bacterial classification (BC) and antibiotic susceptibility testing (AST). Here we demonstrate the use of the fluid handling technique digital microfluidics (DMF) for automated and simultaneous BC and AST using growth metabolic markers. Custom instrumentation was developed for this application including an integrated heating module and a machine-learning-enabled low-cost colour camera for real-time absorbance and fluorescent sample monitoring on multipurpose devices. Antibiotic dilutions along with sample handling, mixing and incubation at 37 °C were all pre-programmed and processed automatically. By monitoring the metabolism of resazurin, resorufin beta-D-glucuronide and resorufin beta-D-galactopyranoside to resorufin, BC and AST were achieved in under 18 h. AST was validated in two uropathogenic E. coli strains with antibiotics ciprofloxacin and nitrofurantoin. BC was performed independently and simultaneously with ciprofloxacin AST for E. coli, K. pneumoniae, P. mirabilis and S. aureus. Finally, a proof-of-concept multiplexed system for breakpoint testing of two antibiotics, as well as E. coli and coliform classification was investigated with a multidrug-resistant E. coli strain. All bacteria were correctly identified, while AST and breakpoint test results were in essential and category agreement with reference methods. These results show the versatility and accuracy of this all-in-one microfluidic system for analysis of bacterial growth and phenotype.

Micromanipulation techniques that are capable of assembling nano/micromaterials into usable structures such as topographical micropatterns (TMPs) have proliferated rapidly in recent years, holding great promise in building artificial electronic and photonic microstructures. Here, a method is reported for forming TMPs based on optoelectronic tweezers in either "bottom-up" or "top-down" modes, combined with in situ photopolymerization to form permanent structures. This work demonstrates that the assembled/cured TMPs can be harvested and transferred to alternate substrates, and illustrates that how permanent conductive traces and capacitive circuits can be formed, paving the way toward applications in microelectronics. The integrated, optical assembly/preservation method described here is accessible, versatile, and applicable for a wide range of materials and structures, suggesting utility for myriad microassembly and microfabrication applications in the future.

There is great interest in the development of micromotors which can convert energy to motion in sub-millimeter dimensions. Micromachines take the micromotor concept a step further, comprising complex systems in which multiple components work in concert to effectively realize complex mechanical tasks. Here we introduce light-driven micromotors and micromachines that rely on optoelectronic tweezers (OET). Using a circular micro-gear as a unit component, we demonstrate a range of new functionalities, including a touchless micro-feed-roller that allows the programming of precise three-dimensional particle trajectories, multi-component micro-gear trains that serve as torque- or velocity-amplifiers, and micro-rack-and-pinion systems that serve as microfluidic valves. These sophisticated systems suggest great potential for complex micromachines in the future, for application in microrobotics, micromanipulation, microfluidics, and beyond., Light-driven micromotors can convert energy to motion in sub-millimeter dimensions. Here, the authors extend this concept and introduce reconfigurable micromachines with multiple components, driven by optoelectronic tweezers, and demonstrate new functionalities.

Background Blood typing, donor compatibility testing, and hematocrit analysis are common tests that are important in many clinical applications, including those found in high-stakes settings such as the trauma center. These tests are typically performed in centralized laboratories with sample batching; the minutes that are lost in this mode can lead to adverse outcomes, especially for critical-care patients. As a step toward providing rapid results at the bedside, we developed a point-of-care hemagglutination system relying on digital microfluidics (DMF) and a unique, automated readout tool, droplet agglutination assessment using digital microfluidics (DAAD). Methods ABO and Rhesus blood grouping, donor crossmatching, and hematocrit assays were developed on a portable DMF platform that allowed for automated sample processing. The result of each assay could be determined by eye or automatically with the DAAD imaging tool. Results DMF-DAAD was applied to 109 samples collected from different sources (including commercial samples, pinpricks from volunteers, and a hospital blood bank), with perfect fidelity to gold-standard results. Some of these tests were carried out by a nonexpert in a hospital trauma center. Proof-of-concept results were also collected from smaller sample sets for donor compatibility testing and hematocrit analysis. Conclusion DMF-DAAD shows promise for delivering rapid, reliable results in a format well suited for a trauma center and other settings where every minute counts.

Microcoil nuclear magnetic resonance (NMR) has been interfaced with digital microfluidics (DMF) and is applied to monitor organic reactions in organic solvents as a proof of concept. DMF permits droplets to be moved and mixed inside the NMR spectrometer to initiate reactions while using sub-microliter volumes of reagent, opening up the potential to follow the reactions of scarce or expensive reagents. By setting up the spectrometer shims on a reagent droplet, data acquisition can be started immediately upon droplet mixing and is only limited by the rate at which NMR data can be collected, allowing the monitoring of fast reactions. Here we report a cyclohexene carbonate hydrolysis in dimethylformamide and a Knoevenagel condensation in methanol/water. This is to our knowledge the first time rapid organic reactions in organic solvents have been monitored by high field DMF-NMR. The study represents a key first step towards larger DMF-NMR arrays that could in future serve as discovery platforms, where computer controlled DMF automates mixing/titration of chemical libraries and NMR is used to study the structures formed and kinetics in real time.

Digital microfluidics (DMF) is a platform that enables highly reconfigurable and automated fluidic operations using a generic device architecture. A unique hallmark of DMF is its "flexibility": a generic device design can be used and reused for many different, divergent fluidic operations. The flexibility of DMF is compromised when devices are permanently modified with embedded sensors. Here we introduce a solution to the "flexibility gap" between fluidic operations in digital microfluidics and embedded sensors: "plug-n-play DMF" (PnP-DMF). In PnP-DMF, devices are designed to allow for rapid and seamless exchange of sensors depending on the application needs. This paper provides "proof of concept" for PnP-DMF using commercial biosensors for glucose and β-ketone, a custom paper-based electrochemical sensor for lactate, and a generic screen-printed electroanalytical cell. We demonstrate that hot-swapping sensors between experiments allows for convenient implementation of complex processes such as automated analysis of blood samples by standard addition. Finally, we explored the suitability for using PnP sensors in tandem with other sensing modalities, combining biosensor-based electrochemical measurement of glucose with a chemiluminescent magnetic bead-based sandwich immunoassay for insulin. The latter is notable, as it constitutes the first report of an analysis of different analytes in both the supernatant and precipitate from a single sample-aliquot in a microfluidic device. The results presented here highlight the versatility of PnP-DMF, illustrating how it may be useful for a wide range of applications in diagnostics and beyond.

Optoelectronic tweezer (OET) is a useful optical micromanipulation technology that has been demonstrated for various applications in electrical engineering and most notably cell selection for biomedical engineering. In this work, we studied the use of light patterns with different shapes and thicknesses to manipulate dielectric microparticles with OET. It was demonstrated that the maximum velocities of the microparticles increase to a peak and then gradually decrease as the light pattern’s thickness increases. Numerical simulations were run to clarify the underlying physical mechanisms, and it was found that the observed phenomenon is due to the co-influence of horizontal and vertical dielectrophoresis forces related to the light pattern’s thickness. Further experiments were run on light patterns with different shapes and objects with different sizes and structures. The experimental results indicate that the physical mechanism elucidated in this research is an important one that applies to different light pattern shapes and different objects, which is useful for enabling users to optimize OET settings for future micromanipulation applications.

Microfluidic sample manipulation is a key enabler in modern chemical biology research. Both discrete droplet-based digital microfluidic (DMF) assays and continuous flow in-channel assays are well established, each featuring unique advantages from the viewpoint of automation and parallelization. However, there are marked differences in the applicable microfabrication materials and methods, which limit the interfacing of DMF sample preparation with in-channel separation systems, such as the gold standard microchip electrophoresis. Simultaneously, there is an increasing demand for low-cost and user-friendly manufacturing techniques to foster the adaptation of microfluidic technology in routine laboratory analyses. This work demonstrates integration of DMF with in-channel separation systems using only low-cost and accessible (non-cleanroom) manufacturing techniques, i.e., inkjet printing of silver for patterning of the driving electrodes and UV curing of off-stoichiometric thiol-ene (OSTE) polymers both for dielectric coating of the electrode arrays and replica molding of the microchannel network. As a dielectric, OSTE performs similar to Parylene C (a gold standard dielectric in electrowetting), whereas its tunable surface and bulk properties facilitate straightforward bonding of the microchannel with the dielectric layer. In addition, a new chip design that facilitates efficient droplet transfer from the DMF part to the microchannel inlet solely by electrowetting is showcased.

Micro-robotics is exploding in popularity, driven by the need to control the position of individual cells and other micronsized particles. There are many examples of optical tweezer-based micro-robots; here we introduce the first microrobotic system that relies on the related technology of optoelectronic tweezers (OET). The optoelectronic micro-robots described here are straightforward to manufacture and can be programmed to carry out sophisticated, multi-axis operations. One particularly useful program is a serial combination of “load,” “transport,” and “deliver,” which can be applied to manipulate a wide range of micron-dimension payloads. Importantly, micro-robots programmed in this manner are much gentler on fragile mammalian cells than conventional OET techniques. The micro-robotic system described here was demonstrated to be useful for single-cell isolation, clonal expansion and RNA sequencing, applications that are becoming increasingly important in the post-CRISPR life-science research landscape. We propose that the OET micro-robotic system, which can be implemented using a microscope and consumer-grade optical projector, will be useful for a wide range of applications in the life sciences and beyond.

A digital microfluidic technique allows for analysis of sub-populations of invading cells by transcriptomics., Microfluidic methods for studying cell invasion can be subdivided into those in which cells invade into free space and those in which cells invade into hydrogels. The former techniques allow straightforward extraction of subpopulations of cells for RNA sequencing, while the latter preserve key aspects of cell interactions with the extracellular matrix (ECM). Here, we introduce “cell invasion in digital microfluidic microgel systems” (CIMMS), which bridges the gap between them, allowing the stratification of cells on the basis of their invasiveness into hydrogels for RNA sequencing. In initial studies with a breast cancer model, 244 genes were found to be differentially expressed between invading and noninvading cells, including genes correlating with ECM-remodeling, chemokine/cytokine receptors, and G protein transducers. These results suggest that CIMMS will be a valuable tool for probing metastasis as well as the many physiological processes that rely on invasion, such as tissue development, repair, and protection.

We check in with past, present, and future Editors-in-Chief, including Andreas Manz (2001–2008), Abe Lee (2017–2020), and Aaron Wheeler (2021–), about the state of the field.

The rectification of ion transport through biological ion channels has attracted much attention and inspired the thriving invention and applications of ionic diodes. However, the development of high-performance ionic diodes is still challenging, and the working mechanisms of ionic diodes constructed by 1D ionic nanochannels have not been fully understood. This work reports the systematic investigation of the design and mechanism of a new type of ionic diode constructed from horizontally aligned multi-walled carbon nanotubes (MWCNTs) with oppositely charged polyelectrolytes decorated at their two entrances. The major design and working parameters of the MWCNT-based ionic diode, including the ion channel size, the driven voltage, the properties of working fluids, and the quantity and length of charge modification, are extensively investigated through numerical simulations and/or experiments. An optimized ionic current rectification (ICR) ratio of 1481.5 is experimentally achieved on the MWCNT-based ionic diode. These results promise potential applications of the MWCNT-based ionic diode in biosensing and biocomputing. As a proof-of-concept, DNA detection and HIV-1 diagnosis is demonstrated on the ionic diode. This work provides a comprehensive understanding of the working principle of the MWCNT-based ionic diodes and will allow rational device design and optimization.

Distance-based detection methods, as used in development of microfluidic paper analytical devices (mu PADs), rely on the dynamic formation of a colored band along the length of the paper microfluidic channels. The color change is driven by the reaction of chromogenic reagents (typically water-insoluble) that are bound to the paper, thus not subject to being washed away by the sample flow along the detection channel. Here, we introduce the use of an anion-exchange filter paper (as a replacement for standard, unmodified filter paper) for distance-based detection in mu PADs, in order to immobilize the water-soluble anionic reagents upon the paper detection channels based on ion-exchange interactions of the oppositely charged paper (protonated tertiary amine groups) and the anionic groups of the reagents. The ion-exchange (IE) paper was initially characterized and its properties were compared with standard cellulose paper. The IE paper was shown to be capable of strong retention of anionic reagents exhibiting acidic functional groups (carboxylic, sulfonic), which become deprotonated and negatively charged when in contact with the IE paper. The effect of the ionic strength (10-250 mM Cl-) and pH (1-13) on the immobilization of the investigated reagents were also determined. The IE-mu PADs were then modified with anionic chromogenic reagents and applied to distance-based determination of total calcium (LOD = 0.03 mM) and total acidity (LOD = 2.5 mM) content in serum and wine samples, respectively. The detailed mechanisms of the developed assays on the IE paper are also discussed. We propose that IE-mu PADs represent a useful new addition to the distance-based detection toolbox and considerably enhance the applicability of such a detection method.

Microrobotics extends the reach of human-controlled machines to submillimeter dimensions. We introduce a microrobot that relies on optoelectronic tweezers (OET) that is straightforward to manufacture, can take nearly any desirable shape or form, and can be programmed to carry out sophisticated, multiaxis operations. One particularly useful program is a serial combination of “load,” “transport,” and “deliver,” which can be applied to manipulate a wide range of micrometer-dimension payloads. Importantly, microrobots programmed in this manner are much gentler on fragile mammalian cells than conventional OET techniques. The microrobotic system described here was demonstrated to be useful for single-cell isolation, clonal expansion, RNA sequencing, manipulation within enclosed systems, controlling cell–cell interactions, and isolating precious microtissues from heterogeneous mixtures. We propose that the optoelectronic microrobotic system, which can be implemented using a microscope and consumer-grade optical projector, will be useful for a wide range of applications in the life sciences and beyond.

The legend in Figure 4a in the previously published version of this article contained erroneous units. The correct units are given in the caption–that is, 1 µM (black), 500 nM (red), 100 nM (blue), 50 nM (cyan), 10 nM (pink), and 0 nM (1× PBS solution, brown).

In digital microfluidics, discrete droplets of fluid are made to move on an open surface with no microchannels. These systems are commonly operated by application of electrical driving forces to an array of electrodes. While these driving forces are well characterized, the dissipative forces opposing droplet movement have not been as thoroughly examined. In recognition of this deficit, we used force-velocity plots to characterize droplet movement in digital microfluidics, which was found to be consistent with a simple theoretical framework for understanding dissipation effects for droplets in two-plate, air-filled devices. Interestingly, in some conditions, a previously unreported ″velocity saturation″ effect was observed. When examined across a range of different liquids, the forces at which this saturation occurs seem to be lower for liquids with smaller surface tensions. Furthermore, when driven at forces that cause saturation, physical phenomena are observed that are akin to what has been reported for stationary droplets in the electrowetting literature. These phenomena are detrimental to device performance, leading to a new "force window" approach that delineates the optimum operation conditions for different liquids. We propose that these findings may be useful for a wide range of applications for experts and new users alike in this growing field.

In this work, we introduce a new type of optical micromanipulation platform that we call patterned optoelectronic tweezers (p-OET). In p-OET devices, the photoconductive layer (that is contiguous in a conventional OET device) is patterned, forming regions in which the electrode layer is locally exposed. We demonstrate that the micro-patterns in the photoconductive layer are useful for repelling unwanted particles/cells, and also for keeping selected particles/cells in place after turning off the light source. We propose that the new technique may be useful for myriad applications in the rapidly growing area of optical micromanipulation.

Immunoprecipitation (IP) is a common method for isolating a targeted protein from a complex sample such as blood, serum, or cell lysate. In particular, IP is often used as the primary means of target purification for the analysis by mass spectrometry of novel biologically derived pharmaceuticals, with particular utility for the identification of molecules bound to a protein target. Unfortunately, IP is a labor-intensive technique, is difficult to perform in parallel, and has limited options for automation. Furthermore, the technique is typically limited to large sample volumes, making the application of IP cleanup to precious samples nearly impossible. In recognition of these challenges, we introduce a method for performing microscale IP using magnetic particles and digital microfluidics (DMF-IP). The new method allows for 80% recovery of model proteins from approximately microliter volumes of serum in a sample-to-answer run time of approximately 25 min. Uniquely, analytes are eluted from these small samples in a format compatible with direct analysis by mass spectrometry. To extend the technique to be useful for large samples, we also developed a macro-to-microscale interface called preconcentration using liquid intake by paper (P-CLIP). This technique allows for efficient analysis of samples100× larger than are typically processed on microfluidic devices. As described herein, DMF-IP and P-CLIP-DMF-IP are rapid, automated, and multiplexed methods that have the potential to reduce the time and effort required for IP sample preparations with applications in the fields of pharmacy, biomarker discovery, and protein biology.

We introduce a method to couple solid-phase microextraction (SPME) with HPLC-MS using digital microfluidics (DMF). In the new system, SPME fibers are used to extract analytes from complex sample solutions, after which the analytes are desorbed into solvent droplets in a DMF device. The open geometry of DMF allows straightforward insertion of SPME fibers without requiring a complicated interface, and automated droplet manipulation enables multiplexed processing of the fibers. In contrast to other multiplexed SPME elution interfaces, the low volumes inherent to DMF allow for pre-concentration of analytes prior to analysis. The new SPME-DMF-HPLC-MS method was applied to the quantification of pg/mL-level free steroid hormones in urine. We propose that this new method will be useful for a wide range of applications requiring cleanup and pre-concentration with convenient coupling to high-performance analytical techniques.

Dopamine (DA) is a classical neurotransmitter and dysfunction in its synaptic handling underlies many neurological disorders, including addiction, depression, and neurodegeneration. A key to understanding DA dysfunction is the accurate measurement of dopamine uptake by dopaminergic neurons. Current methods that allow for the analysis of dopamine uptake rely on standard multiwell-plate based ELISA, or on carbon-fibre microelectrodes used in in vivo recording techniques. The former suffers from challenges associated with automation and analyte degradation, while the latter has low throughput and is not ideal for laboratory screening. In response to these challenges, we introduce a digital microfluidic platform to evaluate dopamine homeostasis in in vitro neuron culture. The method features voltammetric dopamine sensors with limit of detection of 30 nM integrated with cell culture sites for multi-day neuron culture and differentiation. We demonstrate the utility of the new technique for DA uptake assays featuring in-line culture and analysis, with a determination of uptake of approximately ∼32 fmol in 10 min per virtual microwell (each containing ∼200 differentiated SH-SY5Y cells). We propose that future generations of this technique will be useful for drug discovery for neurodegenerative disease as well as for a wide range of applications that would benefit from integrated cell culture and electroanalysis.

Digital microfluidics (DMF) is a droplet-based liquid-handling technology that has recently become popular for cell culture and analysis. In DMF, picoliter- to microliter-sized droplets are manipulated on a planar surface using electric fields, thus enabling software-reconfigurable operations on individual droplets, such as move, merge, split, and dispense from reservoirs. Using this technique, multistep cell-based processes can be carried out using simple and compact instrumentation, making DMF an attractive platform for eventual integration into routine biology workflows. In this review, we summarize the state-of-the-art in DMF cell culture, and describe design considerations, types of DMF cell culture, and cell-based applications of DMF.

We introduce an automated method to facilitate in-line coupling of digital microfluidics (DMF) with HPLC-MS, using a custom, 3D-printed manifold and a custom plugin to the popular open-source control system, DropBot. The method was designed to interface directly with commercial autosamplers (with no prior modification), suggesting that it will be widely accessible for end-users. The system was demonstrated to be compatible with samples dissolved in aqueous buffers and neat methanol and was validated by application to a common steroid-labeling derivatization reaction. We propose that the methods described here will be useful for a wide range of applications, combining the automated sample processing power of DMF with the resolving and analytical capacity of HPLC-MS.

Controlled ion transport through ion channels of cell membranes regulates signal transduction processes in biological systems and has also inspired the thriving development of ionic electronics (ionotronics or iontronics) and biocomputing. However, for constructing highly integrated ionic electronic circuits, the integration of natural membrane‐spanning ion channel proteins or artificial nanomembrane‐based ionic diodes into planar chips is still challenging due to the vertically arranged architecture of conventional nanomembrane‐based artificial ionic diodes. Here, a new design of ionic diode is reported, which allows chip‐scale integration of ionotronics, based on horizontally aligned nanochannels made from multiwalled carbon nanotubes (MWCNTs). The rectification of ion transport through the MWCNT nanochannels is enabled by decoration of oppositely charged polyelectrolytes on the channel entrances. Advanced ionic electronic circuits including ionic logic gates, ionic current rectifiers, and ionic bipolar junction transistors (IBJT) are demonstrated on planar nanofluidic chips by stacking a series of ionic diodes fabricated from the same bundles of MWCNTs. The horizontal arrangement and facile chip‐scale fabrication of the MWCNT ionic diodes may enable new designs of complex but monolithic ionotronic systems. The MWCNT ionic diode may also prove to be an excellent platform for investigation of electrokinetic ion transport in 1D carbon materials.